The Quantikine Human IL-23 Immunoassay is a 4.5 hour solid-phase ELISA designed to measure human IL-23 in cell culture supernates, serum, and plasma. It contains Sf 21-expressed recombinant human IL-23 and has been shown to accurately quantitate the recombinant factor. Results obtained for naturally occurring human IL-23 showed linear curves that were parallel to the standard curves obtained using the kit standards. These results indicate that this kit can be used to determine relative mass values for naturally occurring human IL-23.

Precision

Intra-Assay Precision (Precision within an assay) Three samples of known concentration were tested on one plate to assess intra-assay precision

Inter-Assay Precision (Precision between assays) Three samples of known concentration were tested in separate assays to assess inter-assay precision

Cell Culture Supernates, Serum, EDTA Plasma, Heparin Plasma

Intra-Assay Precision

Inter-Assay Precision

Sample

1

2

3

1

2

3

n

20

20

20

40

40

40

Mean

180

496

1170

203

526

1298

Standard Deviation

12

23.2

49.6

13.3

45.1

78.5

CV%

6.7

4.7

4.2

6.6

8.6

6

Recovery

The recovery of human IL-23 spiked to levels throughout the range of the assay in various matrices was evaluated.

Sample Type

Average % Recovery

Range %

Cell Culture Media (n=4)

93

87-107

EDTA Plasma (n=4)

95

86-106

Heparin Plasma (n=4)

93

85-99

Serum (n=4)

98

88-105

Linearity

To assess the linearity of the assay, samples containing and/or spiked with high concentrations of IL-23 were serially diluted with the Calibrator Diluent to produce samples with values within the dynamic range of the assay.

Store the unopened product at 2 - 8 °C. Do not use past expiration date.

Background: IL-23

IL-23 (Interleukin-23) is a disulfide-linked cytokine composed of a p19 subunit that is unique to IL-23 and a p40 subunit that is shared with IL-12. It is produced by activated macrophages, microglia, and monocyte-derived dendritic cells in response to pathogens. IL-23 induces the earliest recruitment of neutrophils to the site of infection and promotes the development and maintenance of Th17 cells. It also enhances the development of Th17 mediated autoimmunity and tumor progression. IL-23 signals through a receptor complex consisting of IL-12 R beta 1 and IL-23 R. This complex is expressed in mouse Th1 and Th2 cells, bone marrow dendritic cells, activated macrophages and CD4+ CD45Rb(low) memory T cells.

Bring all reagents and samples to room temperature before use. It is recommended that all samples, standards, and controls be assayed in duplicate.

Prepare all reagents, standard dilutions, and samples as directed in the product insert.

Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.

100 µL Assay Diluent

Add 100 µL of Assay Diluent to each well.

100 µL Standard, Control, or Sample

Add 100 µL of Standard, control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours on a horizontal orbital microplate shaker.

Aspirate each well and wash, repeating the process 3 times for a total of 4 washes.

200 µL Conjugate

Add 200 µL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours on the shaker.

Aspirate and wash 4 times.

200 µL Substrate Solution

Add 200 µL Substrate Solution to each well. Incubate at room temperature for 30 minutes on the benchtop. PROTECT FROM LIGHT.

50 µL Stop Solution

Add 50 µL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.

Citations:

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